Construction And Application Of MiRNA Triggered DNA Walkers Induced Photoelectrochemistry Sensor

As a non-protein encoded small molecule nucleotide,miRNA plays an important role in many cellular processes and gene expression.Mi RNA exists in highly stable forms in human serum or plasma,especially in high concentrations in cancer cells and other tumor cells,and becomes a key biomarker for disease diagnosis.Therefore,accurate analysis of miRNA is of great clinical significance.So far,various techniques for detecting miRNA have been developed,among which photoelectric chemical biosensors utilize light sources to excite,and the mode of detecting electrochemical signals has the advantages of low background signals,high sensitivity,and good stability.Due to the extremely low abundance,short length,and high sequence homology of miRNA,it is difficult to accurately quantify miRNA.Therefore,it is necessary to select good photoelectric materials and signal amplification strategies.In this paper,a two-dimensional material,hexagonal boron nitride（h-BN）nanosheet,was utilized and sensitized to improve its photoelectrochemical properties.Combined with the DNA walker signal amplification strategy,a high sensitivity,low detection limit,and high specificity photoelectrochemical biosensor was constructed,providing a strong basis for the diagnosis of diseases.The main research works are as follows:（1）Photochemical biosensor sensitive detection of miRNA-141 based on Cadmium Sulfide（CdS）quantum dots（QDs）anchored h-BN nanosheets and tripod DNA walkers.A highly sensitive photoelectric chemical biosensor based on CdS QDs sensitized porous h-BN nanosheet photoanodes and catalytic hairpin assembly（CHA）formed multipoint tripod DNA walker has been proposed.The h-BN/CdS QDs composite material could promote the transmission of photogenerated electrons and holes,and had excellent photoelectric conversion efficiency.Mi RNA-141 triggered the movement of the tripod DNA walker formed by catalytic hairpin assembly on the DNA functionalized electrode,thereby immobilizing a large amount of alkaline phosphatase（ALP）on the electrode surface,and further catalyzing ascorbic acid 2-phosphate（AAP）in situ to produce ascorbic acid（AA）as an electron donor.Due to the decisive influence of electronic donors on PEC biosensors,sensitive detection of miRNA-141 has been achieved.（2）Assemblable three-dimensional bipedal DNA walker cooperates with CRISPR/Cas12a cascade signal amplification strategy for detecting miRNA-21.Based on the S-BN/CN heterojunction and three-dimensional bipedal DNA walker combined with CRISPR/Cas12a cascade signal amplification technology,a novel photoelectric chemical biosensor was constructed for the sensitive detection of miRNA-21.A heterojunction of S-BN and g-C₃N₄ was constructed by doping sulfur into h-BN,using S-BN/CN as a photosensitive material to provide initial photocurrent,and combining hairpin DNA labeled methylene blue（MB）with a modified electrode to obtain secondary photocurrent amplification.Under the initiation of miRNA-21 and the assistance of Nt.Bsm Al,the bipedal DNA walker outputted an intermediate DNA（O-DNA）.O-DNA combined with cr RNA and was specifically recognized by CRISPR/Cas12a to trigger trans-cleavage.The single-stranded DNA was cut without selectivity,which makes MB away from the surface of the modified electrode and reduced the photocurrent response,thereby realizing the detection of miRNA-21.